Cytotoxic T lymphocytes (CTL) are believed to be exquisitely specific in their ability to recognize and kill antigen-expressing target cells. However, we found that lysis of antigen nonexpressing cells ("innocent bystanders") does take place and is mediated by a Fas ligand (FasL)/Fas-based, but not by a perforin-based, mechanism of cellular cytotoxicity. Thus, the randomly encountered bystanders are killed only if they express sufficient levels of functional Fas and ICAM-1 ligands for LFA-1. These observations are explained by the differential activation thresholds of FasL and/or perforin pathways of cytotoxicity which depend on the intensity of activating stimuli and the nature of cytotoxicity-triggering surface receptor.